The researchers measured the natural amount of thymosin‑beta‑4 (TB4) in horse blood and found it’s about the same no matter the horse’s sex, age, or breed. They also discovered that if blood isn’t separated from cells and is kept at 4 °C, TB4 levels rise quickly because cells break open. Finally, they showed they can spot a synthetic TB4 product in a horse’s plasma after just one dose by looking for a tiny impurity that only shows up in lab‑made versions.

Thymosin <mml:math xmlns:mml="http://www.w3.org/1998/Math/MathML"><mml:mi>&#x3b2;</mml:mi></mml:math> 4 (TB4) is a ubiquitous, highly conserved and abundant peptide among mammals with a critical role in cytoskeleton organization. In spite of its yet non-authorized use as a medicine and being forbidden by the IFHA, the FEI, and the WADA, intelligence and doping control laboratories reported numerous products available online claiming to contain a synthetic acetylated fragment of TB4 or TB4 itself, promoted as a growth factor with regenerative properties. In this article, the first estimation of the endogenous TB4 concentration in racing horses' blood samples was performed through a population study. We reveal that this concentration does not significantly depend on gender, age, nor horse breed. We highlight that the TB4 concentration increases significantly and rapidly in plasma stored at 4&#xb0;C when not separated from blood cells due to cell lysis. Finally, we also demonstrate that the detection of a non-natural synthesis impurity is possible in equine plasma after a single dose administration of a TB4 containing product to a horse.